1. Field of the Invention
This invention is directed to the discovery that binary compositions of an interferon of the alpha, beta or gamma type with tumor necrosis factor, and ternary compositions of an interferon of the alpha and/or beta type with an interferon of the gamma type and tumor necrosis factor have a surprising and unexpected synergistic anti-proliferation effect on neoplastic cells when compared to the activity of the individual components of this invention. The compositions of the present invention are of use in the treatment of neoplasia in animals including humans.
2. Description of the Background Art
A number of proteins occurring naturally in higher organisms are capable of inhibiting the proliferation of tumor cells in vitro and in vivo. These include, inter alia, interferons, lymphotoxins and tumor necrosis factors.
Interferons are divided into three classes differing in their biological and physico-chemical properties. Alpha-interferons (IFN-alpha) include, in humans and a number of other mammals investigated up to now, a family of proteins which are very similar to one another in amino acid sequence and in properties. See, e.g., Stewart, W. E., The Interferon System, Springer Verlag, Wein-New York (1981); Weissmann, C., in Interferon 1981 (Gresser, I., Ed.), pp. 101-154, Academic Press, London (1981); Weck, P. K. et al., J. Gen. Virol., 57:233-237 (1981). Beta-Interferons (IFN-beta) are represented by a single protein in humans and in the majority of other species of mammals investigated. However, a number of IFN-beta subtypes are present in cattle. Wilson, V. et al., J. Mol. Biol., 166:457-475 (1983). Human IFN-beta shows partial sequence homology with human IFN-alpha. Both are relatively acid-stable, and appear to bind to the same membrane receptor. However, human IFN-beta may be differentiated from IFN-alpha, for example, by serological means and by its spectrum of activity on heterologous cells. Stewart, W. E., The Interferon System, Springer Verlag, Wein-New York (1981). Gamma-interferon (IFN-gamma), which is coded for by a single gene in man and in all other species investigated hitherto, is unrelated or only very distantly related to IFN-alpha and IFN-beta in its amino acid sequence. Furthermore, IFN-gamma is acid-unstable and differs clearly from IFN-alpha and IFN-beta in other properties as well. See, Steward, W. E., The Interferon Systems, Springer Verlag, Wein-New York (1981); Epstein, L. in Interferon 1981 (Gresser, I., Ed.), pp. 13-44, Academic Press, London (1981); Gray, P. W. et al., Nature, 295:503-508 (1982); Gray, P. W. & Goeddel, D. V., Proc. Natl. Acad. Sci., USA, 80:5842-5845 (1983).
As a result of these differences, IFN-alpha and IFN-beta are referred to in the literature as type I interferons, while IFN-gamma is referred to as a type II interferon. All these interferons, however, have antiviral activity and a number of immune-modifying activities. Friedman, R. M. and Vogel, S. N., in Advances in Immunology, 34:97-140, Academic Press, London (1983). Type I and type II interferon mixtures show a synergistic activity in various systems. Czarniecky, C. W. et al., J. Virol., 49:490-496 (1984).
Recently, progress in molecular biology has allowed the cloning and production of all classes of human interferons in microorganisms. As a result, it has been possible for the first time to produce sufficiently large quantities of interferons, purified to homogeneity, for the treatment of human diseases in clinical trails. However, the reports of human clinical trails indicate that, in a number of malignant diseases, therapy is unsuccessful or has produced only unsatisfactory results. Moreover, it has been observed that it is often impossible to improve the therapeutic results by increasing the dosage of IFN administered owing to the intolerable side effects which result. Sikora, K. and Smedley, H., Br. Med. J., 286:739-740 (1983); Jones, D. H. et al., Br. J. Cancer, 47:361-366 (1983); Marna, G. et al., J. Biol. Resp. Modif., 2:343-347 (1983).
Tumor necrosis factor (TNF) was originally described as a protein which can be induced in Bacillus Calmette-Guerin-sensitized animals by endotoxin treatment. Carswell, E. A. et al., Proc. Natl. Acad. Sci., USA. 72:3666-3670 (1975). TNF shows cytostatic and/or cytotoxic effects on a number of tumor cell lines in vitro and, in animal models, brings about hemorrhagic necrosis of certain transplantable tumors.
Human B-lymphoblastoid cells lines are known to release a number of lymphokines, including IFN-alpha, macrophage activating factor, migration inhibitory factor and skin reactive factor (Schook, L. B. et al., in Lymphokines: A Forum for Immunoregulatory Cell Products (Pick, E. & Landy, M., Eds.), Vol. 2, pp. 1-19, Academic Press, New York (1981); McEntire, J. E. et al., in Lymphokines and Thymic Hormones: Their Potential Utilization in Cancer Therapeutics (Goldstein, A. L. & Chirigos, M. A., Eds.), pp. 109-119, Raven Press, New York (1981)). Recently, it has been reported that these cells lines also spontaneously produce a protein which, according to various criteria, may be designated human tumor necrosis factor (Hu-TNF). Williamson, B. et al., Proc. Natl. Acad. Sci., USA, 80:5397-5401 (1983). Partially purified culture supernatants of human B-lymphoblastoid cells have shown a synergistic anti-proliferative activity with human IFN-alpha or IFN-gamma. Williamson, B. et al., supra. However, partially purified culture supernatants such as used by Williamson et al. contain many different lymphokines, as noted above, as well as many other potentially unknown impurities. As a result, the true identity of the agent or factor present in these supernatants responsible for the synergistic anti-proliferative activity observed has hitherto been unknown. Thus, the unavailability of a highly purified form of the active agent present in human B-lymphoblastoid cell supernatants has deprived the medical arts of a potentially powerful tool for the treatment of neoplastic disease in man and animals. A need, therefore, has continued to exist to identify the active agent which could then be used in highly purified form along with highly purified human type I and/or type II interferons for disease treatment in man and animals.